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hptA Mutation May Mediate Fosfomycin Resistance in Methicillin-Resistant Staphylococcus aureus Clinical Isolates
Jue Wang1,2, Xiaogang Xu1,2,3, Xiaoyu Zhao1,2
1Institute of Antibiotics, Huashan Hospital, Fudan University, Shanghai, China.
Abstract:
Fosfomycin can be used alone or in combination to treat methicillin-resistant Staphylococcus aureus (MRSA) infection. However, fosfomycin resistance has been observed in MRSA. In S. aureus, fosfomycin resistance is mediated by the fosfomycin-modifying enzyme FosB, or mutations in the target enzyme MurA. Mutations in the chromosomal glpT and uhpT genes, which encode fosfomycin transporters, also result in fosfomycin resistance. The three-component regulatory system HptRSA mediates the expression of uhpT and glpT in S. aureus. This study aimed to investigate the role of hptRSA mutation in fosfomycin resistance in MRSA clinical isolates. We found that hptRSA mutations were common in MRSA strains isolated from our hospital. Most mutations were amino acid substitutions and widely distributed in fosfomycin-sensitive and fosfomycin-resistant strains. However, HptA-truncated mutations were only found in fosB-negative fosfomycin-resistant strains with wild-type uhpT and glpT genes. Quantitative real-time PCR results showed that the transcription level of uhpT decreased by 13.7-25.6-fold in the HptA-truncated strains. Concordantly, the fosfomycin minimum inhibitory concentration (MIC) of HptA-truncated strains was 64-128 μg/mL, while SA240 was 2 μg/mL. The low transcription level of uhpT and high increase in MIC suggest that hptA mutation may lead to fosfomycin resistance in MRSA. We complemented hptA in one of the HptA-truncated clinical strains (SA179), showing reversal of fosfomycin resistance (from 128 to 32 μg/mL). Then we knocked out hptA in S. aureus Newman; fosfomycin MIC increased from 4 to 64 μg/mL, suggesting that HptA mutation may play an important role in fosfomycin resistance.
Insights
Mutations in the HptRSA system, particularly in HptA, contribute to fosfomycin resistance in methicillin-resistant Staphylococcus aureus (MRSA). This resistance is linked to decreased expression of the uhpT gene, impacting fosfomycin transport.
Area of Science:
- Microbiology
- Molecular Biology
- Antimicrobial Resistance
Background:
- Fosfomycin is a crucial antibiotic for treating methicillin-resistant Staphylococcus aureus (MRSA) infections.
- Fosfomycin resistance in MRSA can arise from various mechanisms, including enzymatic modification (FosB), target mutations (MurA), or altered drug transport (GlpT, UhpT).
- The HptRSA system regulates the expression of fosfomycin transporters GlpT and UhpT in S. aureus.
Purpose of the Study:
- To investigate the role of mutations in the HptRSA regulatory system in conferring fosfomycin resistance in clinical MRSA isolates.
- To determine the impact of specific HptRSA mutations on the expression of fosfomycin transporter genes (glpT and uhpT).
- To elucidate the contribution of HptA mutations to fosfomycin resistance mechanisms in MRSA.
Main Methods:
- Screening of clinical MRSA isolates for mutations within the hptRSA operon.
- Analysis of mutation types (e.g., amino acid substitutions, truncations) and their distribution.
- Quantitative real-time PCR to assess the transcriptional levels of uhpT and glpT genes.
- Determination of fosfomycin minimum inhibitory concentrations (MICs).
- Genetic complementation and gene knockout experiments in S. aureus strains.
Main Results:
- Mutations in the hptRSA system were frequently observed in MRSA isolates.
- HptA-truncated mutations were specifically identified in fosfomycin-resistant, fosB-negative MRSA strains with wild-type uhpT and glpT.
- HptA truncation led to a significant decrease in uhpT transcription (13.7-25.6-fold reduction).
- HptA-truncated strains exhibited high fosfomycin MICs (64-128 μg/mL) compared to susceptible strains (2 μg/mL).
- Complementation of hptA restored susceptibility, and hptA knockout increased resistance in a susceptible strain.
Conclusions:
- Mutations in the HptRSA system, particularly HptA truncations, are a significant contributor to fosfomycin resistance in MRSA.
- Reduced uhpT gene expression due to HptA mutations impairs fosfomycin transport, leading to increased resistance.
- The HptRSA system represents a potential therapeutic target for overcoming fosfomycin resistance in MRSA infections.
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